We and others have reported an increased incidence of tumorigenesis in mammary tissue of rodents fed a diet enriched in polyunsaturated fatty acids. We have obtained preliminary evidence that the free fatty acid pool in such tissues is enriched in the hydroxylated metabolite of linoleic acid as a function of linoleic acid in the diet. We are extending this methodological approach based on GC-MS to the analysis of the total lipid pool available in mammary tissue by digesting the tissue in methanolic HCL to transesterify those fatty acids that are tied up as glycerides in phospholipids in the tissue stores. This enhanced approach will give a broader picture of the impact of fatty acid diet on the tissues, but may overwhelm the existing methodology which was developed for the analysis of only the free fatty acid pool. Thus, the important issue in methodology development at this stage is to ensure that our existing methodology has the capacity to deal with the larger analyte load in a reliable manner to support the ongoing studies of tumorigenesis in mammary tissue as a function of fatty acid diet.